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Measurement of homoarginine in human and mouse plasma by LC–MS/MS and ELISA: a comparison and a biological application

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Abstract

Homoarginine (hArg) is a non-essential amino acid that was identified as a risk marker for cardiovascular disease. Several analytical methods have been described for the quantification of hArg in biological samples. The aim of this study was to compare a liquid chromatography–tandem mass spectrometric (LC–MS/MS) approach with a commercially available enzyme-linked immunosorbent assay (ELISA). Determination of hArg concentrations in ELISA calibration standards measured by both methods revealed a correlation coefficient r 2 of 0.99, for LC–MS/MS calibrators r 2 was 0.997. However, linear regression analysis between the two assays for hArg concentrations in human plasma samples revealed a correlation coefficient r 2 of 0.78. Plasma concentrations obtained from LC–MS/MS are on average 29 % higher than those by ELISA. We investigated the hArg-isobaric N ε-trimethyllysine as potential source for the higher observed values, but evaluation of mass spectra indicated that N ε-trimethyllysine did not interfere with hArg quantification in our LC–MS/MS method. Both quantification methods were applied to measure hArg in (1) a case–control study of acute coronary syndrome and (2) l-arginine:glycine amidinotransferase-deficient mice. Our LC–MS/MS and the commercially available ELISA assay are suitable for hArg measurement in human and mouse plasma, but different reference values for each method need to be considered.

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Abbreviations

ACS:

Acute coronary syndrome

AGAT:

L-Arginine:glycine amidinotransferase

ANOVA:

Analysis of variance

CID:

Collision-induced dissociation

CV:

Cardiovascular

ELISA:

Enzyme-linked immunosorbent assay

ESI(+):

Positive electrospray ionization

GC–MS/MS:

Gas chromatography–tandem mass spectrometry

hArg:

Homoarginine

13C6-hArg:

[13C6]-Homoarginine

HPLC:

High-performance liquid chromatography

IQR:

Interquartile range

LC–MS/MS:

Liquid chromatography–tandem mass spectrometry

m/z :

Mass-to-charge ratio

NO:

Nitric oxide

QC:

Quality control

SAP:

Stable angina pectoris

SD:

Standard deviation

SPE:

Solid-phase extraction

UPLC-MS/MS:

Ultrahigh-performance liquid chromatography–tandem mass spectrometry

Wt:

Wild-type

References

  • Araujo P (2009) Key aspects of analytical method validation and linearity evaluation. J Chromatogr B Analyt Technol Biomed Life Sci 877:2224–2234

    Article  CAS  PubMed  Google Scholar 

  • Atzler D, Mieth M, Maas R, Böger RH, Schwedhelm E (2011) Stable isotope dilution assay for liquid chromatography–tandem mass spectrometric determination of L-homoarginine in human plasma. J Chromatogr B Analyt Technol Biomed Life Sci 879:2294–2298

    Article  CAS  PubMed  Google Scholar 

  • Atzler D, Rosenberg M, Anderssohn M, Choe CU, Lutz M, Zugck C, Böger RH, Frey N, Schwedhelm E (2013) Homoarginine—an independent marker of mortality in heart failure. Int J Cardiol 168:4907–4909

    Article  PubMed  Google Scholar 

  • Choe CU, Atzler D, Wild PS, Carter AM, Böger RH, Ojeda F, Simova O, Stockebrand M, Lackner K, Nabuurs C, Marescau B, Streichert T, Müller C, Lüneburg N, De Deyn PP, Benndorf RA, Baldus S, Gerloff C, Blankenberg S, Heerschap A, Grant PJ, Magnus T, Zeller T, Isbrandt D, Schwedhelm E (2013) Homoarginine levels are regulated by l-arginine:glycine amidinotransferase and affect stroke outcome: results from human and murine studies. Circulation 128:1451–1461

    Article  CAS  PubMed  Google Scholar 

  • Davids M, Swieringa E, Palm F, Smith DE, Smulders YM, Scheffer PG, Blom HJ, Teerlink T (2012) Simultaneous determination of asymmetric and symmetric dimethylarginine, L-monomethylarginine, l-arginine, and L-homoarginine in biological samples using stable isotope dilution liquid chromatography tandem mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 900:38–47

    Article  CAS  PubMed  Google Scholar 

  • Di Gangi IM, Chiandetti L, Gucciardi A, Moret V, Naturale M, Giordano G (2010) Simultaneous quantitative determination of N(G), N(G)-dimethyl-l-arginine or asymmetric dimethylarginine and related pathway’s metabolites in biological fluids by ultrahigh-performance liquid chromatography/electrospray ionization-tandem mass spectrometry. Anal Chim Acta 677:140–148

    Article  PubMed  Google Scholar 

  • Hrabák A, Bajor T, Temesi A (1994) Comparison of substrate and inhibitor specificity of arginase and nitric oxide (NO) synthase for arginine analogues and related compounds in murine and rat macrophages. Biochem Biophys Res Commun 198:206–212

    Article  PubMed  Google Scholar 

  • Jones CE, Darcy CJ, Woodberry T, Anstey NM, McNeil YR (2010) HPLC analysis of asymmetric dimethylarginine, symmetric dimethylarginine, homoarginine and arginine in small plasma volumes using a Gemini-NX column at high pH. J Chromatogr B Analyt Technol Biomed Life Sci 878:8–12

    Article  CAS  PubMed  Google Scholar 

  • Kayacelebi AA, Beckmann B, Gutzki FM, Jordan J, Tsikas D (2014) GC-MS and GC–MS/MS measurement of the cardiovascular risk factor homoarginine in biological samples. Amino Acids 46:2205–2217

    Article  CAS  PubMed  Google Scholar 

  • Lehman LJ, Olson AL, Rebouche CJ (1987) Measurement of epsilon-N-trimethyllysine in human blood plasma and urine. Anal Biochem 162:137–142

    Article  CAS  PubMed  Google Scholar 

  • März W, Meinitzer A, Drechsler C, Pilz S, Krane V, Kleber ME, Fischer J, Winkelmann BR, Böhm BO, Ritz E, Wanner C (2010) Homoarginine, cardiovascular risk, and mortality. Circulation 122:967–975

    Article  PubMed  Google Scholar 

  • Meinitzer A, Puchinger M, Winklhofer-Roob BM, Rock E, Ribalta J, Roob JM, Sundl I, Halwachs-Baumann G, März W (2007) Reference values for plasma concentrations of asymmetrical dimethylarginine (ADMA) and other arginine metabolites in men after validation of a chromatographic method. Clin Chim Acta 384:141–148

    Article  CAS  PubMed  Google Scholar 

  • Midttun Ø, Kvalheim G, Ueland PM (2013) High-throughput, low-volume, multianalyte quantification of plasma metabolites related to one-carbon metabolism using HPLC-MS/MS. Anal Bioanal Chem 405:2009–2017

    Article  CAS  PubMed  Google Scholar 

  • Moali C, Boucher JL, Sari MA, Stuehr DJ, Mansuy D (1998) Substrate specificity of NO synthases: detailed comparison of l-arginine, homo-L-arginine, their N omega-hydroxy derivatives, and N omega-hydroxynor-L-arginine. Biochemistry 37:10453–10460

    Article  CAS  PubMed  Google Scholar 

  • Pilz S, Tomaschitz A, Meinitzer A, Drechsler C, Ritz E, Krane V, Wanner C, Böhm BO, März W (2011a) Low serum homoarginine is a novel risk factor for fatal strokes in patients undergoing coronary angiography. Stroke 42:1132–1134

    Article  CAS  PubMed  Google Scholar 

  • Pilz S, Meinitzer A, Tomaschitz A, Drechsler C, Ritz E, Krane V, Wanner C, Boehm BO, März W (2011b) Low homoarginine concentration is a novel risk factor for heart disease. Heart 97:1222–1227

    Article  CAS  PubMed  Google Scholar 

  • Pilz S, Teerlink T, Scheffer PG, Meinitzer A, Rutters F, Tomaschitz A, Drechsler C, Kienreich K, Nijpels G, Stehouwer CD, März W, Dekker JM (2014) Homoarginine and mortality in an older population: the Hoorn study. Eur J Clin Invest 44:200–208

    Article  CAS  PubMed  Google Scholar 

  • Pilz S, Putz-Bankuti C, Meinitzer A, März W, Kienreich K, Stojakovic T, Pieber TR, Stauber RE (2015) Association of homoarginine and methylarginines with liver dysfunction and mortality in chronic liver disease. Amino Acids. doi:10.1007/s00726-015-2000-7

    Google Scholar 

  • Sitar ME, Kayacelebi AA, Beckmann B, Kielstein JT, Tsikas D (2015) Asymmetric dimethylarginine (ADMA) in human blood: effects of extended haemodialysis in the critically ill patient with acute kidney injury, protein binding to human serum albumin and proteolysis by thermolysin. Amino Acids. doi:10.1007/s00726-015-1991-4

    PubMed  Google Scholar 

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Acknowledgments

We thank Mariola Kastner and Anna Steenpaß for their excellent technical support. This study was funded by a grant from the European Commission (Grant 278397 EuRhythDia) and supported by a grant from the DZHK (Grant 81Z1710102). Dr Atzler acknowledges the support of the European Union under a Marie Curie Intra-European Fellowship for Career Development.

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    Authors and Affiliations

    1. Department of Clinical Pharmacology and Toxicology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, 20246, Hamburg, Germany

      Kathrin Cordts, Dorothee Atzler, Vazhma Qaderi, Rainer H. Böger & Edzard Schwedhelm

    2. German Center for Cardiovascular Research (DZHK), Partner Site Hamburg/Kiel/Lübeck, Hamburg, Germany

      Kathrin Cordts, Dorothee Atzler, Rainer H. Böger, Chi-un Choe & Edzard Schwedhelm

    3. Department of Cardiovascular Medicine, University of Oxford, Oxford, UK

      Dorothee Atzler

    4. Department of General and Interventional Cardiology, University Heart Center Hamburg-Eppendorf, Hamburg, Germany

      Vazhma Qaderi & Karsten Sydow

    5. Department of Neurology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

      Chi-un Choe

    Authors
    1. Kathrin Cordts
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    2. Dorothee Atzler
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    5. Rainer H. Böger
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    7. Edzard Schwedhelm
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    Correspondence to Edzard Schwedhelm.

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    The authors declare that they have no conflict of interests.

    Ethical statement

    The study complied with the standards set forth in the Declaration of Helsinki and was approved by the Ethics Committee of the University Medical Center Hamburg-Eppendorf. All patients gave their written informed consent prior to inclusion into the study. The German animal welfare laws for the care and use of animals were followed.

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    E. Schwedhelm and C. Choe contributed equally.

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    Cordts, K., Atzler, D., Qaderi, V. et al. Measurement of homoarginine in human and mouse plasma by LC–MS/MS and ELISA: a comparison and a biological application. Amino Acids 47, 2015–2022 (2015). https://doi.org/10.1007/s00726-015-2037-7

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    • DOI: https://doi.org/10.1007/s00726-015-2037-7

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